Experimental Parasitology

In the past decade, reverse genetics opened new ways to study gene expression in trypanosomes with the development of transient and stable transformation systems. Using stable integrative transformation, it is now possible to carry out precise gene replacements which facilitate the analysis of expression of multicopy gene families (Swindle et al. 1995). Since integration occurs at a low frequency (estimated around 1025), a selectable genetic marker must be used to identify rare transformants. The most common antibiotic resistance genes used for selection of transformed trypanosomes are neor and hygr which confer resistance to G418 and to hygromycin B, respectively. These drugs can be used at concentrations ranging from 1 to 500 mg/ml. Some unusual features of Trypanosoma cruzi, the causative agent of Chagas’ disease, are its presence in a wide variety of hosts and the diversity of symptoms that it causes in human disease. These two facts stimulated studies which revealed a complex biodiversity (Dvorak 1984; Andrade 1990). Our understanding of the parasite variability has benefited greatly from previous population genetics studies: based on multilocus enzyme electrophoresis analysis, T. cruzi populations were shown to have a clonal structure (Tibayrenc et al. 1986). These clonal populations, or clonets, were shown to be divergent for in vitro growth, in vitro metacyclogenesis, and experimental infection in mice (Laurent et al. 1997). Phylogenetic analysis demonstrated that these